The objective of this proposal is to elucidate the ultrastructural and functional features of the mitotic apparatus (MA) in dividing cells. Experiments are designed to gain further insight into the structure and assembly of the MA and its role in chromosome segregation in normal and neoplastic cells. The material to be investigated consists of various cultured mammalian cell lines, including Chinese hamster ovary (CHO) cells, HeLa, mouse BALBc/3T3, and other normal and transformed cell lines as needed. Techniques to be used include conventional electron microscopy, immuno-peroxidase localization of tubulin and actin in the MA, immunofluorescence analysis of spindle and cytoplasmic microtubules, in vitro and in vivo assembly of microtubules, and isolation and characterization of kinetochores from HeLa chromosomes. The areas to be investigated include: (a) further studies on the structure, molecular composition, and function of the kinetochore; (b) further studies of the centriole and its role in the assembly of the MA and chromosome segregation; (c) the comparative sensitivity of cytoplasmic and spindle microtubules to drugs and physical agents; and (d) the structure and assembly of the MA and chromosome movement in somatic cell hybrids.